Repositioning Mifepristone as a Leukaemia Inhibitory Factor Receptor Antagonist for the Treatment of Pancreatic Adenocarcinoma

Abstract

Pancreatic cancer is a leading cause of cancer mortality and is projected to become the second-most common cause of cancer mortality in the next decade. While gene-wide association studies and next generation sequencing analyses have identified molecular patterns and transcriptome profiles with prognostic relevance, therapeutic opportunities remain limited. Among the genes that are upregulated in pancreatic ductal adenocarcinomas (PDAC), the leukaemia inhibitory factor (LIF), a cytokine belonging to IL-6 family, has emerged as potential therapeutic candidate. LIF is aberrantly secreted by tumour cells and promotes tumour progression in pancreatic and other solid tumours through aberrant activation of the LIF receptor (LIFR) and downstream signalling that involves the JAK1/STAT3 pathway. Since there are no LIFR antagonists available for clinical use, we developed an in silico strategy to identify potential LIFR antagonists and drug repositioning with regard to LIFR antagonists. The results of these studies allowed the identification of mifepristone, a progesterone/glucocorticoid antagonist, clinically used in medical abortion, as a potent LIFR antagonist. Computational studies revealed that mifepristone binding partially overlapped the LIFR binding site. LIF and LIFR are expressed by human PDAC tissues and PDAC cell lines, including MIA-PaCa-2 and PANC-1 cells. Exposure of these cell lines to mifepristone reverses cell proliferation, migration and epithelial mesenchymal transition induced by LIF in a concentration-dependent manner. Mifepristone inhibits LIFR signalling and reverses STAT3 phosphorylation induced by LIF. Together, these data support the repositioning of mifepristone as a potential therapeutic agent in the treatment of PDAC.

Discovery of BAR502, as potent steroidal antagonist of leukemia inhibitory factor receptor for the treatment of pancreatic adenocarcinoma

“…To perform STAT3 transactivation, HepG2 (HB, 8065 from ATCC), an immortalized human epatocarcinoma cell line was used, as described previously ( 20 ). On day 0, HepG2 were seeded at 7.5 × 104 cells/well in a 24-well plate and maintained at 37°C and 5% CO2 in E-MEM supplemented with 10% FBS, 1% glutamine, and 1% penicillin/streptomycin.…”

“…The SHAKE algorithm was used to constraint bonds involving hydrogen atoms with two fs integration time steps. Next, the system was minimized and thermally equilibrated as described in our latest work ( 20 ). The MD trajectory was visualized by using Visual Molecular Dynamics (VMD) graphics ver.…”

“…The RNA was extracted from cell lines using and Direct-zol™ RNA MiniPrep w/Zymo-Spin™ IIC Columns (Zymo Research, Irvine, CA, USA)., according to the manufacturer’s protocol as described previously ( 20 ). After purification from genomic DNA by DNase-I treatment (ThermoFisher Scientific, Waltham, MA USA), 2 µg of RNA from each sample was reverse-transcribed using Kit FastGene Scriptase Basic (Nippon Genetics, Mariaweilerstraße, Düren, Germania) in a 20 μL reaction volume.…”

“…MIA PaCa-2 cells were seeded in DMEM complete medium at 800x10 3 cells/well into 24-well plate and used at 70-80% confluence rate. The assay was performed as previously described ( 20 ), particularly on the day 1, the cell monolayers were gently scraped vertically with a new 0.2 mL pipette tip across the centre of the well. After scratching, the well was gently washed twice with PBS (Euroclone, Milan, Italy) to remove the detached cells and cell debris and finally fresh medium containing LIF (10 ng/mL) alone or in combination with BAR502 (10 µM) or EC359 (25 nM) was added into each well.…”

Expression of Selected miRNAs in Normal and Cancer-Associated Fibroblasts and in BxPc3 and MIA PaCa-2 Cell Lines of Pancreatic Ductal Adenocarcinoma

“…Another cytokine of the IL-6 family, leukaemia inhibitory factor (LIF), is also strongly upregulated in PANF, while only insignificantly increased in MELF, Figure 7 . LIF is overexpressed in a variety of solid tumors, including pancreatic tumors [ 92 ], and promotes cancer cell proliferation [ 93 , 94 ]. This gene was found to be induced by hypoxia through HIF1A activation [ 95 ].…”